Department

Department of Veterinary Sciences

First Advisor

Dr. Gerard Andrews

Description

Brucellosis is an infection caused by the bacterium Brucella abortus, which is a concern for both the cattle industry and wildlife organizations. Infection is typically asymptomatic, but has been known to occasionally result in spontaneous abortions in cattle and elk. Humans have also been infected, though treatment is possible. Using the murine (mouse) model, we attempted to characterize and profile the expression of cytokines that occurs during infection with B. abortus Strain 19 (S19), currently a vaccination strain for cattle. Thirty-five mice were vaccinated with S19 and blood samples were collected at seven different time points, with intervals between time points spanning from one week to multiple weeks. Each time point corresponds to five mice which were bled to collect Peripheral Blood Mononuclear Cells (PBMCs). PBMCs were separated by time point and pooled with other PBMCs from mice at that same time point. The pooled PBMC samples were lysed and mRNA was extracted. We utilized Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) to transform mRNA isolated from the PBMCs into cDNA. This cDNA was visualized via DNA gel electrophoresis and analyzed using ImageJ to create ratios of the concentrations of mRNA for cytokines interferon gamma (IFNγ), tumor necrosis factor alpha (TNFα), and interleukin-10 (IL-10) that could be compared to levels of β-actin mRNA (a control). We believe that patterns seen in the expression of these mRNAs may be indicative of the final levels of protein expression of the cytokines we examined. As a result of our work, we observed a pattern of decreasing intensity of the IL-10 response as time from exposure to S19 increased. This suggested the importance of NK cells as mediators of innate immunity in the early stages of S19 infection. We propose that as a result of the maturing immune response, a shift to a more focused attack of the intracellular infection occurs as macrophages and Th1 lymphocytes are allowed to become involved.

Comments

Oral Presentation, Honors Program

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Profiling of expression of IL-10, IFNγ, and TNFα in the PBMCs of mice infected with S19 B. abortus suggests a basic mechanism for the murine immunological response to infection with S19

Brucellosis is an infection caused by the bacterium Brucella abortus, which is a concern for both the cattle industry and wildlife organizations. Infection is typically asymptomatic, but has been known to occasionally result in spontaneous abortions in cattle and elk. Humans have also been infected, though treatment is possible. Using the murine (mouse) model, we attempted to characterize and profile the expression of cytokines that occurs during infection with B. abortus Strain 19 (S19), currently a vaccination strain for cattle. Thirty-five mice were vaccinated with S19 and blood samples were collected at seven different time points, with intervals between time points spanning from one week to multiple weeks. Each time point corresponds to five mice which were bled to collect Peripheral Blood Mononuclear Cells (PBMCs). PBMCs were separated by time point and pooled with other PBMCs from mice at that same time point. The pooled PBMC samples were lysed and mRNA was extracted. We utilized Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) to transform mRNA isolated from the PBMCs into cDNA. This cDNA was visualized via DNA gel electrophoresis and analyzed using ImageJ to create ratios of the concentrations of mRNA for cytokines interferon gamma (IFNγ), tumor necrosis factor alpha (TNFα), and interleukin-10 (IL-10) that could be compared to levels of β-actin mRNA (a control). We believe that patterns seen in the expression of these mRNAs may be indicative of the final levels of protein expression of the cytokines we examined. As a result of our work, we observed a pattern of decreasing intensity of the IL-10 response as time from exposure to S19 increased. This suggested the importance of NK cells as mediators of innate immunity in the early stages of S19 infection. We propose that as a result of the maturing immune response, a shift to a more focused attack of the intracellular infection occurs as macrophages and Th1 lymphocytes are allowed to become involved.